Titanium-Doped Diamond-like Carbon Layers as a Promising Coating for Joint Replacements Supporting Osteogenic Differentiation of Mesenchymal Stem Cells.

Diamond-like carbon (DLC) layers are known for their high corrosion and wear resistance, low friction, and high biocompatibility. However, it is often necessary to dope DLC layers with additional chemical elements to strengthen their adhesion to the substrate. Ti-DLC layers (doped with 0.4, 2.1, 3.7, 6.6, and 12.8 at.% of Ti) were prepared by dual pulsed laser deposition, and pure DLC, glass, and polystyrene (PS) were used as controls. In vitro cell-material interactions were investigated with an emphasis on cell adhesion, proliferation, and osteogenic differentiation. We observed slightly increasing roughness and contact angle and decreasing surface free energy on Ti-DLC layers with increasing Ti content. Three-week biological experiments were performed using adipose tissue-derived stem cells (ADSCs) and bone marrow mesenchymal stem cells (bmMSCs) in vitro. The cell proliferation activity was similar or slightly higher on the Ti-doped materials than on glass and PS. Osteogenic cell differentiation on all materials was proved by collagen and osteocalcin production, ALP activity, and Ca deposition. The bmMSCs exhibited greater initial proliferation potential and an earlier onset of osteogenic differentiation than the ADSCs. The ADSCs showed a slightly higher formation of focal adhesions, higher metabolic activity, and Ca deposition with increasing Ti content.

Hydrogen Sulfide Attenuates Mesenchymal Stem Cell Aging Progress via the Calcineurin-NFAT Signaling Pathway.

Aging is a gradual process that is coupled with a decline in the regenerative capacity of stem cells and a subsequent reduction in tissue function and repair. Hydrogen sulfide (H2S) plays an important role in maintaining the function of stem cells. The present study aimed to investigate the role of H2S in mesenchymal stem cell aging and the underlying mechanism and to provide novel insights into stem cell therapies in elderly people. Bone marrow mesenchymal stem cells (BMMSCs) were isolated from young mice (2 months) and from old mice (12 months). Senescence-associated ß-galactosidase (SA-ß-Gal) activity, reactive oxygen species (ROS) production, ROS scavenging enzymes, and the expression of cell-cycle-related genes were compared between those young and old BMMSCs. The expression of H2S-producing enzymes and the production of H2S in BMMSCs were examined. In vitro osteogenic differentiation and cell senescence were analyzed in young and old BMMSCs before and after H2S treatment. The underlying mechanism was investigated using calcineurin and NFAT1 inhibitors or a Foxp3 siRNA. Bone volume/tissue volume (BV/TV) of femurs in mice was examined using micro-CT with or without systemic injection of an H2S donor. Here, we found that H2S levels in BMMSCs declined with age. When the generation of H2S was blocked with the CBS inhibitor hydroxylamine and the CSE inhibitor dl-propargylglycine, BMMSCs underwent senescence. The elevation of H2S levels rescued BMMSC function in vitro and prevented bone loss in vivo. Mechanistically, H2S represses cell aging via the calcineurin-NFAT1 signaling pathway.

Dissection of Cellular Communication between Human Primary Osteoblasts and Bone Marrow Mesenchymal Stem Cells in Osteoarthritis at Single-Cell Resolution.

Background and Objectives: Osteoblasts are derived from bone marrow mesenchymal stem cells (BMMSCs) and play important role in bone remodeling. While our previous studies have investigated the cell subtypes and heterogeneity in osteoblasts and BMMSCs separately, cell-to-cell communications between osteoblasts and BMMSCs in vivo in humans have not been characterized. The aim of this study was to investigate the cellular communication between human primary osteoblasts and bone marrow mesenchymal stem cells. Methods and Results: To investigate the cell-to-cell communications between osteoblasts and BMMSCs and identify new cell subtypes, we performed a systematic integration analysis with our single-cell RNA sequencing (scRNA-seq) transcriptomes data from BMMSCs and osteoblasts. We successfully identified a novel preosteoblasts subtype which highly expressed ATF3, CCL2, CXCL2 and IRF1. Biological functional annotations of the transcriptomes suggested that the novel preosteoblasts subtype may inhibit osteoblasts differentiation, maintain cells to a less differentiated status and recruit osteoclasts. Ligand-receptor interaction analysis showed strong interaction between mature osteoblasts and BMMSCs. Meanwhile, we found FZD1 was highly expressed in BMMSCs of osteogenic differentiation direction. WIF1 and SFRP4, which were highly expressed in mature osteoblasts were reported to inhibit osteogenic differentiation. We speculated that WIF1 and sFRP4 expressed in mature osteoblasts inhibited the binding of FZD1 to Wnt ligand in BMMSCs, thereby further inhibiting osteogenic differentiation of BMMSCs. Conclusions: Our study provided a more systematic and comprehensive understanding of the heterogeneity of osteogenic cells. At the single cell level, this study provided insights into the cell-to-cell communications between BMMSCs and osteoblasts and mature osteoblasts may mediate negative feedback regulation of osteogenesis process.

Effect of PKH-26-labeled exosomes derived from bone marrow mesenchymal stem cells on corneal epithelium regeneration in diabetic mice.

Background: It is known that bone marrow mesenchymal stem cells (BM-MSCs) could speed up the regeneration of diabetic corneal epithelium. To investigate the effect of exosomes derived from mouse BM-MSCs on corneal epithelium regeneration in diabetic mice. Methods: Diabetic mouse models were established using streptozotocin (STZ), and their central corneal epithelium was scratched under a microscope. The diabetic mice were randomly divided into three groups: the control group was injected with subconjunctival phosphate buffer saline; the exosomes group was treated with a subconjunctival injection of exosomes derived from BM-MSCs; and the BM-MSCs group was treated with a subconjunctival injection of BM-MSCs. The corneal epithelium repair rates in the three groups were compared, and the distribution of the exosomes derived from BM-MSCs labeled with PKH-26 was observed by immunofluorescence. Hematoxylin-eosin staining of the corneal tissue was observed 72 h after the treatments in the three groups. Results: The diabetic mice were successfully established by a blood glucose level >16.7 mmol/L after 8 weeks. The corneal epithelium healing rates in experimental groups 1 and 2 were significantly higher than those of the control group at 24, 48, and 72 h (P<0.05). However, there was no significant difference in the corneal epithelial healing rate between experimental groups 1 and 2 (P>0.05). The exosomes derived from BM-MSCs were found in the superficial corneal stroma in experimental groups 1 and 2, with the majority of the exosomes distributed in the limbal epithelium at the edge of the injury area. The proliferation of corneal epithelial cells in experimental groups 1 and 2 was more obvious than that in the control group. Conclusions: The exosomes derived from BM-MSCs labeled with PKH-26 significantly promoted the repair of corneal epithelial injury in diabetic mice. These exosomes might be a substitute for BM-MSCs in the repair of diabetic keratopathy, suggesting a new idea for the repair of diabetic keratopathy with "cell-free" stem cell therapy, which will require a clinical study.

Regulation of bone and fat balance by Fructus Ligustri Lucidi in ovariectomized mice.

CONTEXT: Fructus Ligustri Lucidi (FLL), a commonly used herb of traditional Chinese medicine (TCM), is the fruit of Ligustrum lucidum Ait. (Oleaceae). The ethanol extract of FLL is a potential candidate for preventing and treating postmenopausal osteoporosis (PMOP) by nourishing the liver and kidneys. OBJECTIVE: This study determines whether an ethanol extract of FLL has anti-osteoporotic effects in ovariectomized (OVX) mice and explores the underlying mechanism. MATERIALS AND METHODS: The OVX model of eight-week-old C57BL/6J female mice was taken, and ovariectomy was used as PMOP. Mice were divided into five groups: sham-operated group (n = 10), OVX group (n = 10), OVX + E2 group (n = 10; 0.039 mg/kg), OVX + FLL group (n = 10; 2 g/kg) and OVX + FLL group (n = 10; 4 g/kg). Mice were treated by gavage with FLL or CMCNa once daily for 8 weeks. We harvested uteri, femur, and tibias from mice; bone mineral density (BMD) and bone microstructure were obtained by X-ray absorptiometry and micro-CT. Furthermore, the effect of FLL on the balance of osteoblast and adipocyte differentiation was investigated using bone marrow mesenchymal stem cells (BMMSCs). RESULTS: The results indicated that FLL did not affect OVX-induced estradiol reduction. Compared with OVX mice, FLL significantly increased BMD (63.54 vs. 61.96), Conn. D (86.46 vs. 57.00), and left tibial strength (13.91 vs. 11.27), decreased Tb. Sp (0.38 vs. 0.44) and body fat content (4.19% vs. 11.24%). FLL decreased osteoclast activity and enhanced RUNX2 expression; inhibited perilipin peroxisome proliferator-activated receptor gamma (PPARγ) expression and adipocyte differentiation from BMMSCs. CONCLUSIONS: FLL prevented additional bone loss and improved bone microstructure in OVX mice by modulating bone and fat balance, suggesting that FLL might be a therapeutic agent for PMOP.

Metformin increases bone marrow adipose tissue by promoting mesenchymal stromal cells apoptosis.

Bone marrow adipose tissue (MAT) has the potential to exert both local and systemic effects on metabolic homeostasis. As a first-line drug used to treat type 2 diabetes mellitus, metformin has conflicting effects on MAT and bone marrow mesenchymal stem cell (BM-MSC) differentiation. Through a series of experiments in vivo and in vitro, we found that except improving the glucose and lipid metabolism disorder in ob/ob mice, 200 mg/kg metformin increased MAT in mice tibia, and prompted osteogenic genes (RunX2, OPN, OCN) and lipogenic genes (Ppar-γ, Cebpα, Scd1) expression in mice bone marrow. However, metformin promoted osteogenesis and inhibited lipogenesis of MSC in vitro, which is inconsistent with the results in vivo. Given MAT being considered the "filler" of the space after the apoptosis of bone marrow stroma, the effect of metformin on MSC apoptosis was examined. We discovered that metformin induces MSC apoptosis in vivo and in vitro. Therefore, we speculated that the increased MAT in mice tibia may be attributed to the filling of adipose tissue after apoptosis of bone marrow stromal cells induced by metformin. The increased MAT may be involved in the regulation of metformin on glucose, lipid, and bone metabolism in diabetic mice, providing a new way to understand the metabolic regulation of metformin. While increased MAT-associated insulin resistance and metabolic disorders may account for the poorer clinical benefits in patients with intensive glucose control.

N-methyl-D-aspartate receptor blockers attenuate bleomycin-induced pulmonary fibrosis by inhibiting endogenous mesenchymal stem cells senescence.

Background: A large number of our previous studies showed that endogenous glutamate and N-methyl-D-aspartate receptor (NMDAR) activation may be involved in various types of acute lung injury, airway inflammation, asthma, and pulmonary fibrosis. In animal models, the transplantation of exogenous bone marrow mesenchymal stem cells (BM-MSCs) is the most promising treatment for idiopathic pulmonary fibrosis. However, there are limited reports on the status of endogenous BM-MSCs in the process of bleomycin-induced pulmonary fibrosis in animals. Methods: We constructed a mouse model of bleomycin-induced pulmonary fibrosis. In vitro, the senescence model of BM-MSCs was constructed with hydrogen peroxide and high concentration of N-methyl-D-aspartate (NDMA). The changes in aging-related indexes were detected by senescence associated beta-galactosidase (SA-ß-gal) staining, western blot, flow cytometry and real time-PCR. The epithelial-mesenchymal transformation (EMT) changes of mouse lung epithelial cells (MLE-12) co-cultured with senescent BM-MSCs were detected by immunofluorescence and western blotting. Results: We observed that endogenous BM-MSCs senescence occurs during bleomycin-induced pulmonary fibrosis in mice, and the model group had a higher expression level of the NMDAR subunit than the control group. We observed a significant increase in NMDAR subunit expression in a hydrogen peroxide-induced senescent cell model in vitro. BM-MSCs showed senescence-related phenotype and cell cycle arrest after high concentration of NMDA treatment. At the same time, the expression levels of the classic Wingless and int-1 (Wnt) pathway protein ß-cantenin and downstream cyclin D1 also changed. In the co-culture of aged BM-MSCs and MLE-12 cells, EMT can be promoted in MLE-12 cells, and MK-801 can partially antagonize the occurrence of EMT. The NMDAR antagonist can partially prevent the above phenomenon. Conclusions: High concentrations of NMDA can promote senescence of BM-MSCs. NMDAR blockers may inhibit endogenous BM-MSCs aging through the WNT signaling pathway, thereby reducing the effect of bleomycin-induced pulmonary fibrosis.

Hypoxia pretreatment improves the therapeutic potential of bone marrow mesenchymal stem cells in hindlimb ischemia via upregulation of NRG-1.

Mesenchymal stem cells (MSCs) are considered a promising treatment for ischemic diseases, but their use is limited due to poor survival after injection. Hypoxia can significantly enhance the survival of MSCs. This study aimed to investigate hypoxia pretreatment of bone marrow mesenchymal stem cells (BM-MSCs) in hindlimb ischemia (HI) and the underlying mechanism. The HI mouse model was established and human BM-MSCs were injected into ischemic skeletal muscles. The blood flow reperfusion and capillary density were measured. In vitro, human BM-MSC cells were treated with hypoxia. The expression of NRG-1 and associated angiogenic factors were measured after knockdown or overexpression of NRG-1. The conditioned medium (CdM) of BM-MSCs was prepared and co-cultured with human umbilical vein endothelial cells (HUVECs), and then, the proliferation, migration, and angiogenesis of HUVECs were detected. After hypoxia pretreatment, NRG-1 expression, clone formation, proliferation, and angiogenic factor secretion from BM-MSCs were increased, while knockdown of NRG-1 reversed these results. In normoxia condition, overexpression of NRG-1 enhanced above factors. Additionally, hypoxia pretreatment of BM-MSCs induced the proliferation and migration of HUVECs and angiogenesis. Moreover, the injection of hypoxia pretreatment of BM-MSCs improved blood reperfusion and capillary density in HI mice, while knockdown of NRG-1 reversed the effect. Furthermore, the PI3K inhibitor and activator reversed the effect of NRG-1 overexpression and knockdown on angiogenesis. We concludes that hypoxia pretreatment of BM-MSCs facilitates angiogenesis and alleviates HI injury via NRG-1/PI3K/AKT pathway.

INTS7-ABCD3 Interaction Stimulates the Proliferation and Osteoblastic Differentiation of Mouse Bone Marrow Mesenchymal Stem Cells by Suppressing Oxidative Stress.

Increased adipocyte and decreased osteoblast differentiation, combined with the ectopic proliferation of bone marrow mesenchymal stem cells (BM-MSCs), represent the primary causes of osteoporosis. The dysregulation of numerous intracellular bioactive factors is responsible for the aberrant differentiation and growth of BM-MSCs. In this study, we focused on a new stimulative factor, integrator complex subunit 7 (INTS7), and its cooperative protein ATP-binding cassette subfamily D member 3 (ABCD3)/high-density lipoprotein-binding protein (HDLBP) in mouse BM-MSCs. We aimed to uncover the effects of the INTS7-ABCD3/HDLBP interaction on BM-MSC biological behaviors and the potential mechanism underlying these effects. Functional in vitro experiments showed that the suppression of the INTS7-ABCD3 interaction rather than HDLBP could impair BM-MSC proliferation and induce cell apoptosis. Moreover, Alizarin Red S and Oil Red O staining, respectively, revealed that INTS7 and ABCD3 knockdown but not HDLBP knockdown could decrease osteoblastic differentiation and accelerate the adipogenic differentiation of BM-MSCs. Mechanistically, reactive oxygen species (ROS) and histone γ-H2AX quantities significantly increased, whereas the levels of antioxidants declined due to INTS7 and ABCD3 inhibition in BM-MSCs. These findings indicated that the suppression of oxidative stress could be involved in the INTS7/ABCD3 co-regulatory mechanisms for BM-MSC proliferation and differentiation, identifying new potential candidates for osteoporosis therapy.

Exosomes secreted by FNDC5-BMMSCs protect myocardial infarction by anti-inflammation and macrophage polarization via NF-κB signaling pathway and Nrf2/HO-1 axis.

BACKGROUND: Exosomes are considered a substitute for stem cell-based therapy for myocardial infarction (MI). FNDC5, a transmembrane protein located in the cytoplasm, plays a crucial role in inflammation diseases and MI repair. Furthermore, our previous study found that FNDC5 pre-conditioning bone marrow-derived mesenchymal stem cells (BMMSCs) could secrete more exosomes, but little was known on MI repair. METHODS: Exosomes isolated from BMMSCs with or without FNDC5-OV were injected into infarcted hearts. Then, cardiomyocytes apoptosis and inflammation responses were detected. Furthermore, exosomes were administrated to RAW264.7 macrophage with LPS treatment to investigate its effect on inflammation and macrophage polarization. RESULTS: Compared with MSCs-Exo, FNDC5-MSCs-Exo had superior therapeutic effects on anti-inflammation and anti-apoptosis, as well as polarizing M2 macrophage in vivo. Meanwhile, the in vitro results also showed that FNDC5-MSCs-Exo decreased pro-inflammatory secretion and increased anti-inflammatory secretion under LPS stimulation, which partly depressed NF-κB signaling pathway and upregulated Nrf2/HO-1 Axis. CONCLUSIONS: FNDC5-BMMSCs-derived exosomes play anti-inflammation effects and promote M2 macrophage polarization via NF-κB signaling pathway and Nrf2/HO-1 Axis, which may develop a promising cell-free therapy for MI.

FNDC5/irisin improves the therapeutic efficacy of bone marrow-derived mesenchymal stem cells for myocardial infarction.

BACKGROUND: The beneficial functions of bone marrow mesenchymal stem cells (BM-MSCs) decline with decreased cell survival, limiting their therapeutic efficacy for myocardial infarction (MI). Irisin, a novel myokine which is cleaved from its precursor fibronectin type III domain-containing protein 5 (FNDC5), is believed to be involved in a cardioprotective effect, but little was known on injured BM-MSCs and MI repair yet. Here, we investigated whether FNDC5 or irisin could improve the low viability of transplanted BM-MSCs and increase their therapeutic efficacy after MI. METHODS: BM-MSCs, isolated from dual-reporter firefly luciferase and enhanced green fluorescent protein positive (Fluc+-eGFP+) transgenic mice, were exposed to normoxic condition and hypoxic stress for 12 h, 24 h, and 48 h, respectively. In addition, BM-MSCs were treated with irisin (20 nmol/L) and overexpression of FNDC5 (FNDC5-OV) in serum deprivation (H/SD) injury. Furthermore, BM-MSCs were engrafted into infarcted hearts with or without FNDC5-OV. RESULTS: Hypoxic stress contributed to increased apoptosis, decreased cell viability, and paracrine effects of BM-MSCs while irisin or FNDC5-OV alleviated these injuries. Longitudinal in vivo bioluminescence imaging and immunofluorescence results illustrated that BM-MSCs with overexpression of FNDC5 treatment (FNDC5-MSCs) improved the survival of transplanted BM-MSCs, which ameliorated the increased apoptosis and decreased angiogenesis of BM-MSCs in vivo. Interestingly, FNDC5-OV elevated the secretion of exosomes in BM-MSCs. Furthermore, FNDC5-MSC therapy significantly reduced fibrosis and alleviated injured heart function. CONCLUSIONS: The present study indicated that irisin or FNDC5 improved BM-MSC engraftment and paracrine effects in infarcted hearts, which might provide a potential therapeutic target for MI.

Identification and Comparison of piRNA Expression Profiles of Exosomes Derived from Human Stem Cells from the Apical Papilla and Bone Marrow Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are multifunctional stem cells that exist in almost all human tissues. In addition to their self-renewal and multidirectional differentiation potential, they also have valuable immunomodulatory abilities. Bone marrow mesenchymal stem cells (BMMSCs) are the first discovered MSCs and are the most widely studied. Stem cells from the apical papilla (SCAP) are derived from the apical papilla of incompletely developed teeth and play an important role in the formation and development of tooth root. Recent studies have shown that mesenchymal stem cell-derived exosomes (MSC-exo) have similar biological functions as MSCs. Moreover, increasing evidence has highlighted the functional relationship between noncoding regulatory RNAs, especially microRNAs, and MSC-exo. However, few studies have addressed the role of PIWI-interacting RNAs (piRNAs) in MSC-exo. To develop a better understanding of the biological functions of SCAP and BMMSCs, we compared and analyzed the piRNA expression profiles of the exosomes derived from human SCAP (SCAP-exo) and the exosomes of BMMSCs (BMMSC-exo). A total of 593 and 920 known piRNAs were identified from SCAP-exo and BMMSC-exo, respectively, and 21 piRNAs were found to be differentially expressed. In addition, we predicted the target genes of the differentially expressed piRNAs, and the target genes were subjected to the Gene Ontology enrichment and the Kyoto Encyclopedia of Genes and Genomes pathway analysis, revealing the possible biological functions of these differentially expressed piRNAs. We found that the target genes of the differentially expressed piRNAs mainly involved in biological regulation, cellular processes, metabolic processes, binding, and catalytic activity, which are closely related to the biological functions of MSCs. In conclusion, this study confirmed the differential expression profiles of piRNAs in SCAP-exo and BMMSC-exo and provided useful insights for further study of their functions.

Calvarial bone defects in ovariectomised rats treated with mesenchymal stem cells and demineralised freeze-dried bone allografts.

BACKGROUND: The aim of the study was to investigate the ability of a combination of bone marrow mesenchymal stem cells (BM-MSCs) with and without demineralised freeze-dried bone allografts (DFDBAs) to induce bone regeneration in calvarial defects in ovariectomised rats. MATERIALS AND METHODS: Critical size defects were filled with a combination of DFDBAs and BM-MSCs or BM-MSCs alone. Eight weeks after calvarial surgery, the rats were sacrificed. The samples were analysed histologically and immunohistochemically. RESULTS: No difference was observed in vascularisation between groups C1 (animals with cranial defect only, control group) and O1 (animals with cranial defect only, ovariectomy group). Intramembranous ossification was observed at a limited level in groups C2 (animals with cranial defect with MSCs, control group) and O2 (animals with cranial defect with MSCs, ovariectomy group) compared to C1 and O1. In group C3 (animals with DFDBAs with MSCs, control group), the fibrous structures of the matrix became compact as a result of a bone graft having been placed in the cavity, but in group O3 (animals with DFDBAs with MSCs, ovariectomy group), the fibrous tissue was poorly distributed between the bone grafts for the most parts. CONCLUSIONS: We conclude that the insertion of BM-MSCs enhances bone healing; however, the DFDBA/BM-MSC combination has little effect on overcoming impaired bone formation in ovariectomised rats.

Increased Expression of FGF-21 Negatively Affects Bone Homeostasis in Dystrophin/Utrophin Double Knockout Mice.

Duchenne muscular dystrophy (DMD) is the most common muscular dystrophy seen in children. In addition to skeletal muscle, DMD also has a significant impact on bone. The pathogenesis of bone abnormalities in DMD is still unknown. Recently, we have identified a novel bone-regulating cytokine, fibroblast growth factor-21 (FGF-21), which is dramatically upregulated in skeletal muscles from DMD animal models. We hypothesize that muscle-derived FGF-21 negatively affects bone homeostasis in DMD. Dystrophin/utrophin double-knockout (dKO) mice were used in this study. We found that the levels of circulating FGF-21 were significantly higher in dKO mice than in age-matched WT controls. Further tests on FGF-21 expressing tissues revealed that both FGF-21 mRNA and protein expression were dramatically upregulated in dystrophic skeletal muscles, whereas FGF-21 mRNA expression was downregulated in liver and white adipose tissue (WAT) compared to WT controls. Neutralization of circulating FGF-21 by i.p. injection of anti-FGF-21 antibody significantly alleviated progressive bone loss in weight-bearing (vertebra, femur, and tibia) and non-weight bearing bones (parietal bones) in dKO mice. We also found that FGF-21 directly promoted RANKL-induced osteoclastogenesis from bone marrow macrophages (BMMs), as well as promoted adipogenesis while concomitantly inhibiting osteogenesis of bone marrow mesenchymal stem cells (BMMSCs). Furthermore, fibroblast growth factor receptors (FGFRs) and co-receptor ß-klotho (KLB) were expressed in bone cells (BMM-derived osteoclasts and BMMSCs) and bone tissues. KLB knockdown by small interfering RNAs (siRNAs) significantly inhibited the effects of FGF21 on osteoclast formation of BMMs and on adipogenic differentiation of BMMSCs, indicating that FGF-21 may directly affect dystrophic bone via the FGFRs-ß-klotho complex. In conclusion, this study shows that dystrophic skeletal muscles express and secrete significant levels of FGF-21, which negatively regulates bone homeostasis and represents an important pathological factor for the development of bone abnormalities in DMD. The current study highlights the importance of muscle/bone cross-talk via muscle-derived factors (myokines) in the pathogenesis of bone abnormalities in DMD. © 2019 American Society for Bone and Mineral Research.

SRC3 expressed in bone marrow mesenchymal stem cells promotes the development of multiple myeloma.

SRC3 plays critical roles in various biological processes of diseases, including proliferation, apoptosis, migration, and cell cycle arrest. However, the effect of SRC3 expression in mesenchymal stem cells (MSCs) on multiple myeloma (MM) is not clear yet. In our study, MSCs (MSC-SRC3, MSC-SRC3-/-) and MM cells were co-cultured in a direct or indirect way. The proliferation of MM cells was studied by CCK-8 and colony formation assays. The apoptosis and cell cycle of MM cells were detected by flow cytometry. In addition, the expressions of proteins in MM cells were detected by western blot analysis and the secretions of cytokines were measured by ELISA. Our data showed that the expression of SRC3 in bone marrow mesenchymal stem cells (BM-MSCs) could promote cell proliferation and colony formation of MM cells through accelerating the transformation of the G1/S phase, no matter what kind of culture method was adopted. Meanwhile, SRC3 expressed in BM-MSCs could inhibit the apoptosis of MM cells through the caspase apoptosis pathway and mitochondrial apoptosis pathway. Moreover, SRC3 could enhance the adhesion ability of MM cells through up-regulating the expression of adhesion molecules including CXCL4, ICAM1, VLA4, and syndecan-1. SRC3 also played a regulatory role in the progress of MM through the NF-κB and PI-3K/Akt pathways. SRC3 expressed in MSCs was found to promote the growth and survival of MM cells, while SRC3 silencing in MSCs could inhibit the development of MM. These results would be useful for developing a more effective new strategy for MM treatment.

Inositol pyrophosphates mediated the apoptosis induced by hypoxic injury in bone marrow-derived mesenchymal stem cells by autophagy.

OBJECTIVE: To investigate the potential effect of IP7 on the autophagy and apoptosis of bone marrow mesenchymal stem cells (BM-MSCs) caused by hypoxia. METHODS: BM-MSCs isolated from adult male C57BL/6 mice were exposed to normoxic condition and hypoxic stress for 6 h, 12 h, and 24 h, respectively. Then, flow cytometry detected the characteristics of BM-MSCs. Furthermore, N6-(p-nitrobenzyl) purine (TNP) was administrated to inhibit inositol pyrophosphates (IP7). TUNEL assay determined the apoptosis in BM-MSCs with hypoxia. Meanwhile, RFP-GFP-LC3 plasmid transfection and transmission microscope was used for measuring autophagy. In addition, Western blotting assay evaluated protein expressions. RESULTS: Hypoxic injury increased the autophagy and apoptosis of BM-MSCs. At the same time, hypoxic injury enhanced the production of IP7. Moreover, hypoxia decreased the activation of Akt/mTOR signaling pathway. At last, TNP (inhibitor of IP7) repressed the increased autophagy and apoptosis of BM-MSCs under hypoxia. CONCLUSION: The present study indicated that hypoxia increased autophagy and apoptosis via IP7-mediated Akt/mTOR signaling pathway of BM-MSCs. It may provide a new potential therapy target for myocardial infarction (MI).

Androgen attenuates antitumor effects of gastric cancer cells by bone marrow mesenchymal stem cells via restricting the JNK signaling activation.

BACKGROUND: Researches on bone marrow mesenchymal stem cells (BMMSCs) have generated controversial results in tumor research. In the present study, we aimed to explore the functions of BMMSCs on gastric cancer and the possible mechanism in a mimicking microenvironment of the stomach. METHODS: Transwell co-cultured system was used to co-culture BMMSCs and gastric cancer SGC-7901 cells. In some experiments, androgen and its antagonist were added into the cells as required. Cell viability, cell apoptosis, mRNA and protein expressions of apoptosis- and JNK signaling- associated genes were respectively determined by performing cell counting kit-8, flow cytometry, quantitative real-time PCR and western blot. RESULTS: Androgen contributed to the growth of BMMSCs and SGC-7901 cells. In co-cultured system, BMMSCs not only suppressed SGC-7901 cell viability, induced cell apoptosis and promoted tumor necrosis factor (TNF)-α release, but also regulated the level of Bax/Bcl-2 and elevated the expressions of phosphorylation (p)-JNK and p53. After adding androgens, the anti-tumor effects of BMMSCs were weakened. Meanwhile, the antagonists of androgens could partially recover BMMSCs in vitro inhibitory effects on gastric cancer cells by activation of JNK signaling. CONCLUSIONS: This study demonstrated the important roles of BMMSCs on the growth and apoptosis of gastric cancer cells in vitro. Additionally, in the mimicking microenvironment of the stomach, androgen weakened the antitumor effects of BMMSCs by limiting JNK signaling activation, suggesting that androgen antagonist may be a promising adjuvant drug to BMMSCs in gastric cancer therapy.

Autologous platelet-rich gel combined with in vitro amplification of bone marrow mesenchymal stem cell transplantation to treat the diabetic foot ulcer: a case report.

The diabetic foot ulcer (DFU) is the leading cause of the high mortality and morbidity rates of diabetes patients, and the DFU accounts for approximately 15% of all diagnosed diabetes cases in China. Traditional treatment is typically ineffective for DFUs. Here, we present a case of DFU that was successfully treated with an autologous platelet-rich gel (APG) and in vitro amplification of bone marrow mesenchymal stem cell (BMMSC) transplantation. A 54-year-old woman initially presented with a right foot diabetic ulcer at the hospital. A wound at the lateral malleolus of the right foot was observed with exudation and infection. The standard treatment included glucose reduction with insulin, blood lipid control with atorvastatin, circulation improvement with alprostadil, anti-infection treatment with sensitive antibiotics, debridement, dressing, and continuous negative pressure suction, and after the standard treatment, the APG combined with in vitro amplification of BMMSC transplantation was used to help the healing of the ulcer. All of the above interventions may have contributed to the healing of the ulcer, and an APG combined with in vitro amplification of BMMSCs may promote DFU healing. The difficulty of DFU treatment remains a challenge, particularly in diabetic patients who develop foot ulcers, due to the complexity of its multifaceted pathogenesis. This case represents an effective adjuvant treatment for such patients.

Long non-coding RNA H19 mediates mechanical tension-induced osteogenesis of bone marrow mesenchymal stem cells via FAK by sponging miR-138.

Bone marrow mesenchymal stem cells (BMMSCs) provide the biological basis for bone reconstruction. Mechanical tension stimulation as a potent modulator is able to promote osteogenic capability of BMMSCs. Long non-coding RNAs (LncRNAs) as competing endogenous RNAs (ceRNAs) for microRNAs, are postulated to regulate the osteogenic differentiation of stem cells. However, the mechanism how (whether) lncRNAs mediates tension-induced osteogenesis of BMSCs still remains poor understood. Here, human BMMSCs (hBMMSCs) were subjected to mechanical tension (10%, 0.5Hz). Results showed that mechanical tension could enhance osteogenic differentiation and increase H19 expression. H19 deficiency suppressed tension-induced osteogenic differentiation, demonstrating that H19 could mediate tension-induced osteogenesis in hBMMSCs. Besides, mechanical tension could suppress miR-138 expression, and down-regulated miR-138 promoted tension-induced osteogenesis in hBMMSCs. Luciferase reporter assays illustrated that H19 had binding sites with miR-138, and H19 deficiency increased miR-138 level, demonstrating that H19 may act as a ceRNA for miR-138 in hBMMSCs. Luciferase reporter assays also showed that miR-138 could target PTK2,a gene encoding focal adhesion kinase (FAK). Up-regulated miR-138 impaired increased FAK expression induced by mechanical tension. The relationship among H19, miR-138 and FAK under tension condition was further studied. H19 deficiency inhibited FAK expression, which could be partly rescued by knock-downing miR-138. In addition, suppressed tension-induced osteogenic differentiation in H19 defective cells was partly rescued by miR-138 knockdown. Taken together, this study indicated that H19 is a positive regulator in tension-induced osteogenesis of hBMMSCs through acting as a ceRNA for miR-138 and then up-regulating downstream FAK.

Into the eyes of bone marrow-derived mesenchymal stem cells therapy for myocardial infarction and other diseases.

Applications of bone marrow-derived mesenchymal stem cells (BM-MSCs) have been documented for diseases occur in the sports system, the central nervous system, the cardiovascular system etc. However, poor viability of donor stem cells after transplantation limits their therapeutic efficiency. Although the autophagy theory has been reported, the underlying mechanisms are still poorly understood. Isolation and culture methods of mesenchymal stem cells are currently concentrate on four ways. Overall, BM-MSCs have both important research significance and clinical application value in cell replacement therapy, gene therapy and reconstruction of tissues as well as organs especially for myocardial infarction (MI). In this article, we review the biological characteristics of BM-MSCs and its research progress especially in MI.